The human malaria parasite, Plasmodium falciparum, is transmitted by a mosquito vector, which consumes infected red blood cells from one individual, and after sexual development and sporogony of the parasite, injects infectious sporozoites into another individual. Not all blood stage parasites are competent to develop in the mosquito. Asexual blood stage parasites can only complete their life cycle in the human host, giving rise to more parasites and the clinical symptoms of malaria. Other blood stage parasites differentiate sexually into male and female gametocytes that can continue developing only in the mosquito. Expression of unique gametocyte stage genes is necessary for sexual differentiation and successful mosquito transmission. The long term objective of this research proposal is to produce a physical map of the stage specific transcriptional activity along large continuous stretches of DNA (tens to hundreds of kB) flanking two model gametocyte stage genes, the pfs16 and pfg27/25 genes. These genes are transcribed exclusively in the sexual stages. In laboratory isolates which have lost the capacity to differentiate sexually and complete their life cycle, the pfs16 and pfg27/25 genes are not transcribed. I propose to isolate these genes on yeast artificial chromosomes (YACs) to test the hypothesis that sexual stage genes are coordinately transcribed. Asexual and gametocyte stage cDNA or RNA probes will identify and determine the stage specificity of new blood stage transcription units on the YAC clones around the model gametocyte stage genes. This unique approach may identify new gametocyte stage antigens as potential targets for transmission blocking immunity. We have identified a mutant P. falciparum isolate in which the pfg27/25 gene is aberrantly transcribed in the asexual stages. Comparison of the transcriptional environment around the gene in wild type asexual parasites and in the mutant will determine whether inappropriate activation of a gene modifies the transcriptional stage of nearby genes.